Infections due to extracellular bacteria continue to pose a significant global health problem. This is due in large part to the continual emergence of antibiotic-resistant strains. Hence, there exists an urgent need for development of protective vaccines. Immunity is mediated by antibodies to the bacterial polysaccharides (PS) as well as proteins. However, little is known regarding the parameters that mediate in vivo anti-PS and anti-protein responses to intact extracellular bacteria, although such information has relevance to the rational design of immunotherapies for these agents. We have established an in vivo model system for investigating the mechanism of induction of anti-PS and anti-protein Ig isotypes in response to intact Streptococcus pneumoniae. Specifically, the Ig isotype response to the phosphorylcholine (PC) determinant, present on the bacterial cell wall C-PS is studied and compared to the humoral response to a cell wall protein, pneumococcal surface protein A (PspA). We show that induction of optimal anti-PC and anti-PspA responses both require CD4+TCR-a/b+ T cells and B7-dependent costimulation, although memory fails to develop for induction of PC-specific Ig. Of interest, the mechanisms underlying the T cell-dependence of these two responses are distinct. We further show that dendritic cells (DCs) can phagocytose S. pneumoniae upon transfer into naive mice, induce both anti-PC and anti-PspA Ig responses, and the formation of PspA-specific memory. The general aims of this application are to elucidate the mechanisms by which DCs respond to and process an intact extracellular bacterium for induction of both T cell-dependent PC- and PspA-specific Ig isotypes in vivo, and determine the mechanisms underlying the distinct forms of T cell help that stimulate these respective antigen-specific Ig isotype responses. Specifically, we will utilize a number of in vitro and in vivo model systems to determine 1) the parameters that regulate DC activation and antigen presentation in response to R36A, 2) the relative contribution of DC subsets, and 3) the role of DC cytokines and accessory molecules, including CD40, MHC class 11, and Toll-like receptors. In this context, 4) the differential requirements for DC stimulation of T cell help for the anti-PC versus the anti-PspA response will be determined. These data will be the first to establish the detailed parameters that mediate a physiological antigen-specific humoral immune response to an intact extracellular bacterium, including the delineation of the fundamental differences between polysaccharide and protein-specific Ig isotype responses.